Oral infection represents an important route of human immunodeficiency virus (HIV) and human cytomegalovirus (HCMV) transmission, leading to global consequences. HCMV infection is among the most common causes of oral diseases associated with AIDS patients. Furthermore, HIV and HCMV are found in oral tissues and their interaction is believed to play an important role for their successful oral transmission as well as for the development of viral-induced oral diseases. However, little is currently known about the mechanism of HCMV and HIV oral infections, partially because these infections are primarily limited to human cells, and few cultured tissue or animal models for studying HCMV and HTV infections in oral mucosa have been reported. Recent in vitro studies indicate that productive HIV infection occurs in normal human oral keratinocytes and is enhanced in the presence of environmental factors in oral mucosa, such as alcohol, which up-regulates CXCR4 receptor expression. Our preliminary studies suggest that HCMV productively infects cultured human gingival and buccal tissues and significantly enhances HIV infection in cultured oral mucosa. The overall objective of this proposed research is to develop human gingival and buccal tissue culture model and use these cultured tissues to study the infection of HCMV and HIV and their interactions in oral mucosa. In the initial part of the proposed study, in vitro human buccal and gingival tissue culture models will be developed. Tissues will then be infected with a library of HCMV mutants with deletion of a single viral open reading frame. Viral mutants that are defective in growth in cultured tissues will be identified and characterized. Furthermore, the function of HCMV determinants essential for viral oral infection will be studied. Finally, tissues will be infected with HIV in the absence and presence of different HCMV mutants, and the mechanism of how HIV oral infection is affected by HCMV will be investigated. The proposed research will lead to the development of new tissue culture models to study HIV and HCMV infection in oral mucosa. These studies will also identify viral genes required for HCMV replication in oral mucosa and investigate the mechanism of HIV-HCMV interactions in oral cavity. The results will provide insight into the mechanism of HCMV and HIV infection in oral mucosa and facilitate the development of novel strategies for treatment and prevention of the transmission as well as infections of these viruses in oral cavity.